SUMMARY It is largely unknown how early life stress exposure triggers neural circuit dysfunction and leads to neuropsychological disorders. While genetic model organisms, such as the nematode C. elegans, are instrumental in determining genetic and environmental factors that affect development, one current major bottleneck is the lack of appropriate instrumentation to image cellular activities while recording behavior in juveniles, because of difficulties associated with their small size. The overall objective in this project is to address this lack of technology by developing such an engineered system and apply it to study the impact of early exposure to stress. The overall objectives will be attained by pursuing two specific aims. 1) To develop a droplet- based platform for imaging the behavior and neuronal calcium transients in larvae. Droplet microfluidics offers unique features for partitioning and precise control of micro-carriers, and thus can deliver chemicals with precise timing and dosage. In addition, droplets are of the appropriate size to larvae and will therefore address the challenge of efficiently manipulating small larvae and stimulating them. 2) Using the multi-modal imaging platform, to probe neuronal dynamics during development upon adverse stimulation in the case of olfactory imprinting. One will measure the activity of neurons involved in memory formation as well as neurons involved in memory retrieval in nave and imprinted animals throughout development. This will serve as a testbed for the technology development, as well as gaining insights into the biological process. The research proposed here is innovative: first, it will develop original techniques using droplet microfluidics to manipulate small animals and produce chemical gradients in droplets that were previously impossible; second, it will allow for performing functional neuronal imaging on multicellular organisms encapsulated in droplets. Because a droplet system presents key advantages for serial processing, such a platform paves the way for high-throughput screening based on neuronal activity. The research is significant because it resolves a problem in handling very small and fragile animals, and addressing this problem will greatly facilitate not only the study of imprinting but also the study of any developmental processes that occur in juveniles. The technology will move forward drugs screens for developmental disorders.